Why do some proteins in a family prove very hard to target, while others bind a whole list of inhibitors? This paper takes a look at a particularly dramatic example in the kinase field. That’s a good place for studying such things, since there are a lot of kinases out there, and a lot of kinase inhibitors. Many of those compounds have been profiled across some part of the “kinome” to look at their selectivity – and believe me, some of them (even or maybe even especially approved drugs) are blunderbusses. But this analysis flips the question around, building on this 2017 work towards establishing a standardized data set for kinase inhibition, and asks about the apparent specificities of the enzymes themselves for small-molecule inhibition.
In general, across 398 enzymes you find a mean of about 17 inhibitors for each one in the data set. At one end of the scale, there are kinases out there that have only one known inhibitor. This might partly be due to lack of clinical interest, but remember, this is the result of taking all sorts of known kinase inhibitors and running them across enzymes that they were never targeted for, so that flattens out the bias to a good degree. And at the other end, there is a cluster of eight kinase enzymes that bind a mean of 99 of the known inhibitors (!), across a whole range of structures. Some of these in fact (like YSK4 and DDR1) have not been the targets of many drug discovery efforts, which helps rule out discovery bias as well.
What’s different about them? Phylogenetically, the eight enzymes are not particularly related in any evolutionary sense. X-ray crystal structures provided what looks like the answer, though. As kinase fans know, there are two major binding modes for such compounds, “DFG-in” and “DFG-out”, referring to a particular conserved stretch of Asp-Phe-Gly. There are successful kinase inhibitors in both binding category; the DFG-in conformation is the catalytically active one under normal conditions, and has traditionally been considered the more stable one. What’s odd is that these promiscuous enzymes, when exposed to classic DFG-in binding inhibitors, bind them in the DFG-out mode instead. It appears that these proteins have a particular salt bridge elsewhere in their structure that stabilizes the DFG-out conformation, and this opens them up to a huge range of potential binders. (That might well also explain the low catalytic activity displayed by many of the group, too).
The comparison between the classic “kinome tree”, which is based on sequence homology and the differences in inhibitor promiscuity are interesting. Anyone who’s worked in the field will recognize the shape of the tree at right; it’s been with us since the canonical 2002 kinome paper. But you can see from the overlay of this current work that the salt-bridged DFG-out enzymes (large green circles) are scattered through the tree in ways that the standard sequence analysis would not have made clear. This particular structural motif would appear to have emerged more than once – you should never assume that evolutionary history, as revealed by sequence, is going to tell you a straightforward flowing narrative.
Why some of these kinases have adopted what seems to be a deliberately less active lifestyle, with the DFG-out form stabilized, is open for speculation. The way that this conformation opens them up to a wider range of small-molecule inhibitors is probably a side effect; kinases aren’t regulated in the cell through endogenous small molecules that bind to the active site in this way. And it’s important to remember that this doesn’t mean that you can’t find a selective inhibitors of (say) DDR1. You most certainly can. What this work tells you, though, is that finding starting points for such chemical matter is going to be relatively easy.